The pituitary gland is critical for growth, reproduction and maintenance of homeostatic physiological mechanisms in mammals. The development of the pituitary is dependent upon the actions of an array of transcription factors, including the LHX3 LIM-homeodomain protein, to establish the hormone-producing cells of the anterior lobe. Mutations in the genes encoding several of these essential transcription factors result in severe pediatric pituitary diseases such as combined pituitary hormone deficiency (CPHD) disease. These diseases can have severe impacts on the development and quality of life of affected children, and the underlying etiologies and progression of the diseases are unknown in most cases. The proposed research project and training plan will use a novel mouse model (W227Ter knock-in) of a severe form of pediatric CPHD caused by a mutation in the LHX3 gene that results in production of a protein lacking the carboxyl terminus. In order to understand the molecular and cellular basis for the onset of this CPHD disease, pituitaries of W227Ter knock-in mouse embryos will be examined at multiple stages of embryonic development for expression of the characteristic anterior pituitary hormones, cellular differentiation markers, and critical transcription factors. This analysis is not possible in the human patients and will therefore provide important insights into the nature of the human disease. Recently, a niche of adult stem cells expressing transcription factors such as LHX3 has been shown to exist along the edge of the anterior pituitary. A gradual depletion of pituitary function over time in CPHD patients may be related to the loss of stem/precursor cells. Therefore, both embryonic and adult pituitaries will be examined for expression of stem cell markers. These studies will increase the understanding of the development of these debilitating diseases and therefore contribute to improved diagnosis and treatment. This three-year training plan will be a research intensive yet comprehensive plan for Dr. Prince. The new techniques learned such as cryosectioning, immunohistochemistry, and in situ hybridization, as well as the focused study on pituitary development will complement her existing extensive research experience. Dr. Rhodes' laboratory facilities, worldwide collaborations and commitment to excellent mentoring will add to the successful completion of the proposed project as well as overall career development. Participating in career development workshops, the Preparing Future Faculty program, presenting at and attending departmental seminars and national meetings, and interacting with other scientists in a collaborative environment will ensure a comprehensive training and career development experience. This will position Dr. Prince to obtain a position as a biomedical researcher and educator at a four-year institution, thereby meeting her career goal. PUBLIC HEALTH RELEVANCE: This research will allow molecular characterization of the symptoms and progression of a severe form of a pediatric hormone deficiency disease. This type of analysis is not possible in the patients. The findings will contribute to improved diagnosis and treatment of these diseases.